Revealing the molecular structure of unpurified silk and other biomaterials

Breaking new ground 5. mar 2023 3 min Postdoctoral Fellow Giulia Giubertoni Written by Kristian Sjøgren

Researchers used advanced polarised two-dimensional infrared spectroscopy to investigate the molecular structure of unpurified silk. The study demonstrates that the molecular properties of the building blocks of pure biomaterials can be studied without harsh chemical treatment that can alter these molecular properties.

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Researchers, engineers and companies all over the world want to understand how many of nature’s materials work because they often function much better than human-made materials. Improving understanding of molecular structures can help researchers and industry to recreate similar biomaterials in laboratory flasks or large production tanks.

For example, spider silk is one of the strongest known materials, is flexible and can adapt to the surroundings by changing character when it gets wet. So far, however, how spider silk achieves this is unknown.

New research shows how to use polarised two-dimensional infrared spectroscopy directly on a biomaterial to reveal the intrinsic protein structures underlying its mechanical properties.

“All biomaterials from bones to spider silk need specific mechanical properties to fulfil their biological role. For example, both skin and spider silk must be elastic so that they do not crack when deformed,” explains a researcher involved in the study, Giulia Giubertoni, Postdoctoral Fellow, Faculty of Science, Van ‘t Hoff Institute for Molecular Sciences, University of Amsterdam, Netherlands.

Giulia Giubertoni collaborated with Sander Woutersen, Professor of Physical Chemistry, Faculty of Science, University of Amsterdam.

The research, which has been published in Biomacromolecules, is the result of a collaboration between the University of Amsterdam, the University of Sheffield and Aarhus University.

Biomaterials lose their special mechanical properties

Spider silk and other biomaterials can potentially become huge industrial ventures, but this requires making spider silk artificially, since harvesting it from living spiders is nearly impossible. This necessitates determining the molecular structure of the underlying building blocks, typically proteins, and how changes in temperature or humidity or being deformed changes the structure.

The proteins comprising the biomaterial determine its mechanical properties, and errors in the protein sequence will eliminate the special mechanical properties and its biological function.

“This study showed how a special technology can provide greater insight into the protein structure of a biomaterial without processing it. It enables researchers to understand how the protein structure changes when the biomaterial adapts its mechanical properties to changes in the environment,” says Giulia Giubertoni.

Improved infrared spectroscopy

Infrared spectroscopy is frequently used to study the molecular properties of biomaterials’ building blocks. The technology excels for studying the functional biomaterials on which Giulia Giubertoni focuses, but the molecular information on all aspects of the protein structure and mechanical properties is often not detailed enough for full insight.

The researchers instead used a different technology to study unpurified silk.

Silk is interesting to study because researchers can examine it without processing it, such as dissolving it. This proof-of-concept study analysed pure biomaterials without any kind of chemical processing.

Spider webs in the forest

The researchers used polarised two-dimensional infrared spectroscopy to study the silk, which is more complicated than the more conventional infrared spectroscopy.

However, two-dimensional infrared spectroscopy often provides much more information about the molecular details because it is sensitive to vibrational couplings in proteins, providing detailed structural information.

“Two-dimensional infrared spectroscopy enables us to obtain more molecular information on the building blocks of biomaterials and how changes in the environment affect the molecular structure of the biomaterial,” explains Giulia Giubertoni.

Because the researchers do not have to pretreat the biomaterial before studying it, they can investigate how it functions under different conditions.

For example, a spider web in the forest must be stable and functional in the midday sun but also in the dewy early morning and in rain, when the web must bear the weight of the water.

Biomaterials adapt to their environment

The mechanical properties in the protein structure enable the spider web to handle both scenarios, because the spider silk can change its protein structure and adapt to the different conditions.

Since the researchers do not have to dissolve a biomaterial before analysing it with two-dimensional infrared spectroscopy, they can also simulate different conditions and determine how the biomaterial behaves depending on the environment.

“This is the very big advantage of being able to carry out our studies on pure biomaterials,” says Giulia Giubertoni.

Greater insight into different biomaterials will enable them to be created in the laboratory or enable researchers to understand what happens when the biomaterials do not function properly.

Learning about brittle-bone disease

The next step in Giulia Giubertoni’s research is to learn more about collagen and why it does not work properly in various diseases. The Dutch Research Council (NWO) is supporting this part of the research.

For example, a single mutation in the proteins in collagen causes osteogenesis imperfecta (brittle-bone disease), and the bones of the people with osteogenesis imperfecta are fragile and break easily without specific cause.

“Making better materials inspired by nature or understanding mechanical failure in various diseases requires understanding how the molecular and macroscopic properties of samples that are nearly pure biomaterials are linked. This study showed that polarised two-dimensional infrared spectroscopy combined with other technologies might help us to obtain this insight,” concludes Giulia Giubertoni.

In situ identification of secondary structures in unpurified Bombyx mori silk fibrils using polarized two-dimensional infrared spectroscopy” has been published in Biomacromolecules. The Novo Nordisk Foundation awarded grants to co-author Tobias Weidner for the projects Nanosurface Scattering Sum Frequency Spectrometer – NanoScat (2018) and Deciphering the Role of Atmospheric Microbial Aerosols (DRAMA) (2019).

Giubertoni is a post-doctoral fellow at the Van't Hoff Institute for Molecular Sciences of the University of Amsterdam. Together with Prof.S.Woutersen...

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